The overall objective of this research proposal is to utilize recombinant DNA technology to identify and isolate the major immunogens of Treponema pallidum (Nichols), and to eventually test these immunogens as possible effective vaccines against experimental syphillis in rabbits. These studies may also provide fundamental knowledge about the genetic characteristics of the organism and the evolution of the humoral immune response during the progression of the disease. An E. coli-T. pallidum hybrid plasmid clone bank, comprised of more than 53,000 recombinant DNA clones and representing the entire T. pallidum (Nichols) genome, has been constructed in this laboratory. Bacterial clones will be screened for the presence of T. pallidum DNA inserts and the absence of contaminating rabbit DNA by the methods of colony hybridization and Southern gel transfers, both using nick translated radioactive T. pallidum DNA and rabbit DNA as hybridization probes. Inserts of T. pallidum DNA will be isolated from the total clone bank plasmid DNA and used to determine the analytical complexity (size) of the T. pallidum (Nichols) genome. Analytical complexity experiments will involve the construction of DNA-DNA hybridization data curves following free-solution Cot analyses of the hybridizing DNA components. Bacterial clones synthesizing specific T. pallidum immunogens as plasmid gene translation products will be identified from the clone bank by solid-phase radioimmunoassay techniques employing protective IgG antibody isolated from hyperimmune rabbit serum. Immunogens from the bacterial clones will be identified, characterized, and isolated from bacterial clones using analytical and preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis in conjection with immunoprecipitation. Antibody to trephonemal immunogens will be produced in rabbits, purified, and used to assess the different antibody specificities to T. pallidum. The ability of antibodies raised against the purified immunogens to inactivate virulent treponemes will also be examined. Purified treponemal immunogens will ultrimately be tested for efficacy as potential vaccinogens against syphilitic infection in rabbits.